DESCRIPTION: K+ channels regulate resting potential, spike spacing and many other cellular processes, failure of which leads to diseases found in heritabl syndromes. K+ channels have been found in microbes; the one in yeast has been cloned and analyzed under patch clamp. While many aspects of channel molecular biology have been elucidated through site-directed mutagenesis, further structure-function studies can be advanced through the use of microbial genetics. Mutant selection and biophysical analysis have recently been married in the study of the yeast K+ channel, YKC1. This union yielded thus far, among others, mutations in regions that destabilize close-state conformations as deduced from the gating kinetics of K+ currents. The continued application of power of yeast genetics and biophysical analysis will be pursued. The approach allows specific alteration of the YKC1 gene in plasmid without modification of the yeast genome. As in progress, more gain-of-function mutants will be isolated to exhaustively map the sites involved in gating between closed and open conformations. The window of selection will be altered to select mutants of different phenotypes. Other YKC1-channel mutants with specific ion sensitivity or over-tolerance and those that rescue a K+-uptake deficiency will be isolated to further dissect the ion filter and the rectification mechanism. In all instances, the sequence defects will be correlated with the electrophysiological defects detected by patch-clamping native yeast cells as well as Xenopus oocytes expressing the YKC1 genes. Molecular structure-and -function correlations gathered will be used to address existing models or to construct new models of this K+ channel.